During the assembly of neural circuits, newly born neurons migrate to specific locations and extend processes in stereotyped manners to contact the appropriate synaptic partners. This is most evident in the vertebrate retina which is organized into three nuclear layers separated by two intervening synaptic plexiform layers. Despite the fundamental nature of this organization little is known about the cellular mechanisms that position neuronal cell types and direct dendrite extension. This study examines the function of the atypical cadherin protein Fat3 which coordinates these events during amacrine cell development. Fat3 is an unusually large cadherin molecule with a mass of ~500Kd that contains 34 extracellular cadherin domains and is present throughout the developing inner plexiform layer (IPL). Analysis of knockout mice reveals that fat3 is necessary for directing the polarized extension of amacrine cell dendrites into the IPL as well as properly distributing amacrine cells between the inner nuclear layer (INL) and the ganglion cell layer (GCL). In the absence of fat3 ectopic amacrine cell dendrites elaborate outside of the IPL resulting in the formation of two additional synaptic layers. In this project the molecular and cellular mechanism(s) of Fat3 function will be determined using novel lines of fat3 knockout and conditional knockout mice. Specifically transgenic reporters will be used to identify morphological classes of amacrine and ganglion cells expressing Fat3 and distinguish between a general function for amacrine cell development versus a function in the assembly of specific neuronal circuits. Additional in vitro experiments will dissect the regulatory mechanisms that control Fat3 signaling including alternative splicing and dynamic interactions with different cytoplasmic proteins. Finally ganglion cell development will be analyzed to determine if Fat3 is also required for the morphogenesis and distribution of this cell type, and central projections will be examined to determine if Fat3 is necessary for axonal development. Although focused on the function of Fat3 in the retina, the proposed work will nevertheless provide important insight into the role of Fat cadherins during neurodevelopment and in other systems where they are highly expressed such as the inner ear and kidney. PUBLIC HEALTH RELEVANCE: In the vertebrate retina the organization of neurons and synaptic connections into circuits connecting the light detecting photoreceptors with retinal ganglion cells that project to the brain is essential for the formation of visual images. This project is designed to determine how retinal neurons are positioned in the retina and extend dendrites to contact the correct synaptic partners. By understanding the developmental events that build retinal circuitry we can predict how these circuits are altered in disease states, identify targets for pharmacological intervention, and contribute to our understanding of the biological basis of vision.